Much has been learned in recent years about the central mechanisms controlling the initiation of mammalian puberty, it is now clear that no isolated pathway or cellular subset is responsible for this control, which instead appears to be exerted by regulatory gene networks. A global approach for the system-level identification of such networks has never been attempted, essentially due to the lack of appropriate technology, and the relative paucity of genetic and biochemical details that can be assimilated into a testable biological model. The emergence of high-throughout approaches and novel computational methods for data analysis is giving us for the first time the opportunity of identifying genetic modules involved in the hierarchical control of puberty. Using some of these approaches we have singled out a group of genes that may represent the first identified genetic network involved in the neuroendocrine control of female puberty. These genes share the feature of having been earlier identified as involved in "tumor suppression". In this application, prepared in response to PA Number PA-03-079, we propose to combine emerging technology to develop the novel concept that "tumor suppressor genes" (TSGs) form a functionally interactive network that - operating within neuronal and glial subsets of the hypothalamus - provide the system-wide control underlying the pubertal activation of LHRH secretion. The following aims are proposed: 1. To construct a fist-stage architectural model of the TSG genetic network, and identify the cellular sites of expression of both upper echelon genes and genes situated at nodal points in the network. To achieve the first part of the aim we will test novel computational methods not yet applied to the dissection of complex biological processes in mammals; the second part will be accomplished using immunohistochemistry-/n situ hybridization procedures. 2. To experimentally test the viability of the TSG network via perturbation of in silico predicted key components of the system. We propose to achieve this aim by disrupting TSG expression using the technology of small-interfering RNAs (siRNA) delivered to the hypothalamus via the novel approach of modified lentiviruses carrying dual siRNA transcriptional cassettes. Although siRNAs are widely used to perturb gene expression in mammalian cells, the methodology to achieve long-term silencing effects in vivo is still in its infancy. [unreadable] [unreadable]